Maintained by Robin Tecon, microbiologist and postdoctoral researcher at the Swiss Federal Institute of Technology Zürich. This blog is about bacteria (and other microbes) and the scientists who study them.

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Sunday, July 29, 2012

Indole teaches persistence to bacteria

When a bacterial infection is treated with antibiotics, bacteria
that are in a so-called dormant, inactive state may escape death – this because
antibiotics only kill growing
bacteria. It becomes a serious problem when these sleeping beauties start to
grow again, in particular when they do so after the period of antibiotic
treatment has ended… Thus, an infection that was apparently cured could be
followed by a secondary infection days or weeks later. This problematic phenomenon
is called bacterial persistence, and it should not be confused with bacterial
resistance, in which growing bacteria are immune to one or several antibiotics.

Now what about indole? (The molecule displayed on top of
this post.) Actually indole is present in very common and important biomolecules,
such as the amino acid tryptophan, the animal hormone serotonin and the plant growth
hormone auxin. We have known for more than a century that E. coli produces indole in stationary phase (Lee, 2010), and it
does so thanks to an enzyme called tryptophanase, which cleaves tryptophane
into indole, pyruvate and ammonia.

But E.
coli is not the only bacterium capable of that: more than 85 species (both
Gram-negative and Gram-positive) can synthesize indole (Lee, 2010). For a long time the biological functions of
indole were overlooked, but now we know that indole can act as an extracellular
signal and can for instance increase antibiotic resistance and control biofilm
formation in E. coli.

The group of Jim Collins, at Boston University, noted that
indole was produced under conditions (stationary phase, limited nutrients)
conducive to bacterial persistence. They decided to investigate the possible
role of indole signaling in the formation of bacterial peristers in E. coli, and they published their
results in May 2012 as a brief communication in Nature Chemical Biology (NicoleVega et al., 2012).

And what they show clearly is that exposure to indole increases
the appearance of persisters by a ten-fold factor in a population of E. coli! In addition, they showed that
bacteria that were not producing tryptophanase (thus incapable of producing
indole from tryptophane) formed much less persister cells.

How does the
signaling work? Well, it is not fully understood yet, but Vega et al. showed that indole is acting
extracellularly, because E. coli
cells that are missing a specific indole transporter (Mtr, which imports indole inside
the bacterium) are more likely to become persistent than normal E. coli cells.

Curiously, even though the bacteria are exposed to a fixed
concentration of indole in a liquid environment, they will not all react the
same way: it is clear that the population’s response to indole is heterogeneous.
Vega demonstrated this by using a fluorescent reporter system that was
responsive to indole signaling. Thus, they could follow the expression of
fluorescence in individual cells and study the differences among the
population. After antibiotic treatment, the persisters were also the ones that
glowed most in response to indole!

Finally, the group of Jim Collins looked at the
transcriptome of E.coli during indole
signaling, and they found that two pathways, the so-called phage-shock response
and oxidative stress response were stimulated by indole. Now if you disrupt
both phage-shock and oxidative stress pathways in E. coli, the induction of persistence by indole disappears… Interestingly, they also found that cells exposed to indole
had no higher expression in drug export, which is required for bacterial
resistance. Hence, it confirms that bacterial persistence, not resistance, is
at play here.

Model for indole signaling and persister formation in E. coli, from Vega et al.,
2012. Some cells in the population turn into persisters (orange
cells) after exposure to indole. I thank Jim Collins for the permission
to reproduce the figure here.

But this phenomenon may not be totally specific to indole,
since Vega et al. showed that treatment with hydrogen peroxide could also activate the
oxidative stress pathway and lead to persister formation.

The authors conclude as follows:

“The bacterial signaling molecule indole is sensed in a
heterogeneous manner by a population of cells, causing induction of OxyR and
phage-shock pathways via a periplasmic or membrane component, thereby inducing
the creation of a persistent subpopulation. Indole is not toxic at
physiological concentrations, but it triggers protective responses, acting to
inoculate a subpopulation (persisters) against possible future stress.”

What I would be curious to know is what is responsible for the
heterogeneous response? The authors did not address this question (maybe in a
future work?) but acknowledged its importance:

“These findings add to an understanding of persister
formation as bacterial ‘bet-hedging’ strategy in uncertain environments.”

Indeed, it can be valuable for bacteria to maintain diversity within the population, in particular when their life conditions are susceptible to change often. The more you look at it, the more you find such examples of phenotypic variabilityin microbial ecology. It definitely plays a major role in the bacterial world.

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